Endothelin receptor antagonists

ABSTRACT

Pyrrolo[2,3-b]pyridine derivatives are described which are endothelin receptor antagonists.

This application is a 371 of PCT/US95/07220 filed Jun. 7, 1995 which isa continuation of U.S. application Ser. No. 08/255,622 filed Jun. 9,1994, now abandoned.

FIELD OF THE INVENTION

The present invention relates to novel pyrrolopyridine derivatives,pharmaceutical compositions containing these compounds and their use asendothelin receptor antagonists.

Endothelin (ET) is a highly potent vasoconstrictor peptide synthesizedand released by the vascular endothelium. Endothelin exists as threeisoforms, ET-1, ET-2 and ET-3. [Unless otherwise stated "endothelin"shall mean any or all of the isoforms of endothelin]. Endothelin hasprofound effects on the cardiovascular system, and in particular, thecoronary, renal and cerebral circulation. Elevated or abnormal releaseof endothelin is associated with smooth muscle contraction which isinvolved in the pathogenesis of cardiovascular, cerebrovascular,respiratory and renal pathophysiology. Elevated levels of endothelinhave been reported in plasma from patients with essential hypertension,acute myocardial infarction, subarchnoid hemorrhage, atherosclerosis,and patients with uraemia undergoing dialysis.

In vivo, endothelin has pronounced effects on blood pressure and cardiacoutput. An intravenous bolus injection of ET (0.1 to 3 nmol/kg) in ratscauses a transient, dose-related depressor response (lasting 0.5 to 2minutes) followed by a sustained, dose-dependent rise in arterial bloodpressure which can remain elevated for 2 to 3 hours following dosing.Doses above 3 nmol/kg in a rat often prove fatal.

Endothelin appears to produce a preferential effect in the renalvascular bed. It produces a marked, long-lasting decrease in renal bloodflow, accompanied by a significant decrease in GFR, urine volume,urinary sodium and potassium excretion. Endothelin produces a sustainedantinatriuretic effect, despite significant elevations in atrialnatriuretic peptide. Endothelin also stimulates plasma renin activity.These findings suggest that ET is involved in the regulation of renalfunction and is involved in a variety of renal disorders including acuterenal failure, cyclosporine nephrotoxicity, radio contrast induced renalfailure and chronic renal failure.

Studies have shown that in vivo, the cerebral vasculature is highlysensitive to both the vasodilator and vasoconstrictor effects ofendothelin. Therefore, ET may be an important mediator of cerebralvasospasm, a frequent and often fatal consequence of subarachnoidhemorrhage.

ET also exhibits direct central nervous system effects such as severeapnea and ischemic lesions which suggests that ET may contribute to thedevelopment of cerebral infarcts and neuronal death.

ET has also been implicated in myocardial ischemia (Nichols et al, Br.J. Pharm. 99:597-601, 1989 and Clozel anc Clozel, Circ. Res., 65:1193-1200, 1989) coronary vasospasm (Fukauda et al., Eur. J. Pharm. 165:301-304, 1989 and Luscher, Circ, 83: 701, 1991) heart failure,proliferation of vascular smooth muscle cells, (Takagi, Biochem &Biophys. Res. Commun.; 168: 537-543, 1990, Bobek et al., Am. J. Physiol.258:408-C415, 1990) and atherosclerosis, (Nakaki et al., Biochem. &Biophys. Res. Commun. 158: 880-881, 1989, and Lerman et al., New Eng. J.of Med, 325: 997-1001, 1991). Increased levels of endothelin have beenshown after coronary balloon angioplasty (Kadel et al., No. 2491 Circ.82: 627, 1990).

Further, endothelin has been found to be a potent constrictor ofisolated mammalian airway tissue including human bronchus (Uchida etal., Eur J. of Pharm. 154: 227-228 1988, LaGente, Clin. Exp. Allergy 20:343-348, 1990; and Springall et al., Lancet, 337: 697-701, 1991).Endothelin may play a role in the pathogenesis of interstitial pulmonaryfibrosis and associated plumonary hypertension, Glard et al., ThirdInternational Conference on Endothelin, 1993, p. 34 and ARDS (AdultRespiratory Distress Syndrome), Sanai et al., Supra, p. 112.

Endothelin has been associated with the induction of hemorrhagic andnecrotic damage in the gastric mucosa (Whittle et al., Br. J. Pharm. 95:1011-1013, 1988); Raynaud's phenomenon, Cinniniello et al., Lancet 337:114-115, 1991); Crohn's Disease and ulcerative colitis, Munch et al.,Lancet, Vol. 339, p. 381; Migraine (Edmeads, Headache, February 1991 p127); Sepsis (Weitzberg et al., Circ. Shock 33: 222-227, 1991; Pittet etal., Ann. Surg. 213: 262-264, 1991), Cyclosporin-induced renal failureor hypertension (Eur. J. Pharmacol., 180: 191-192, 1990, Kidney Int, 37:1487-1491, 1990) and endotoxin shock and other endotoxin induceddiseases (Biochem. Biophys. Res. Commun., 161: 1220-1227, 1989, ActaPhysiol. Scand. 137: 317-318, 1989) and inflammatory skin diseases.(Clin Res. 41:451 and 484, 1993).

Endothelin has also been implicated in preclampsia of pregnancy. Clarket al., Am. J. Obstet. Gynecol. March 1992, p. 962-968; Kamor et al., N.Eng. J. of Med., Nov. 22, 1990, p. 1486-1487; Dekker et al., Eur J. Ob.and Gyn. and Rep. Bio. 40 (1991) 215-220; Schiff et al., Am. J. Ostet.Gynecol. February 1992, p. 624-628; diabetes mellitus, Takahashi et al.,Diabetologia (1990) 33:306-310; and acute vascular rejection followingkidney transplant, Watchinger et al., Transplantation Vol. 52, No. 4,pp. 743-746.

Endothelin stimulates both bone resorption and anabolism and may have arole in the coupling of bone remodeling. Tatrai et al, Endocrinology,Vol. 131, p. 603-607.

Endothelin has been reported to stimulate the transport of sperm in theuterine cavity, Casey et al., J. Clin. Endo and Metabolism, Vol. 74, No.1, p. 223-225, therefore endothelin antagonists may be useful as malecontraceptives. Endothelin modulates the ovarian/menstrual cycle,Kenegsberg, J. of Clin. Endo. and Met., Vol. 74, No. 1, p. 12, and mayalso play a role in the regulation of penile vascular tone in man, Lauet al, Asia Pacific J. of Pharm, 1991, 6:287-292 and Tejada et al., J.Amer. Physio. Soc. 1991, H1078-H1085. Endothelin also mediates a potentcontraction of human prostatic smooth muscle, Langenstroer et al., J.Urology, Vol. 149, p. 495-499.

Thus, endothelin receptor antagonists would offer a unique approachtoward the pharmacotherapy of hypertension, renal failure, ischemiainduced renal failure, sepsis-endotoxin induced renal failure,prophylaxis and/or treatment of radio-contrast induced renal failure,acute and chronic cyclosporin induced renal failure, cerebrovasculardisease, myocardial ischemia, angina, heart failure, asthma, pulmonaryhypertension, pulmonary hypertension secondary to intrinsic pulmonarydisease, atherosclerosis, Raynaud's phenomenon, ulcers, sepsis,migraine, glaucoma, endotoxin shock, endotoxin induced multiple organfailure or disseminated intravascular coagulation, cyclosporin-inducedrenal failure and as an adjunct in angioplasty for prevention ofrestenosis, diabetes, preclampsia of pregnancy, bone remodeling, kidneytransplant, male contraceptives, infertility and priaprism and benignprostatic hypertrophy.

SUMMARY OF THE INVENTION

This invention comprises pyrrolopyridine derivatives represented byFormula (Ia-Id) and pharmaceutical compositions containing thesecompounds, and their use as endothelin receptor antagonists which areuseful in the treatment of a variety of cardiovascular and renaldiseases including but not limited to: hypertension, acute and chronicrenal failure, cyclosporine induced nephrotoxicity, stroke,cerebrovascular vasopasm, myocardial ischemia, angina, heart failure,benign prostratic hypertrophy, migraine, pulmonary hypertension,atherosclerosis, and as an adjunct in angioplasty for prevention ofrestenosis.

This invention further constitutes a method of treatment of diseasescaused by an excess of endothelin, which comprises administering to ananimal in need thereof an effective amount of a compound of Formula(Ia-Id).

DETAILED DESCRIPTION OF THE INVENTION

The compounds of this invention are represented by structural Formula(Ia-Id): ##STR1## wherein

R₁ is --X(CH₂)_(n) Ar or --X(CH₂)_(n) R₈ or ##STR2##

R₂ is hydrogen, Ar or (c);

P₁ is --X(CH₂)_(n) R₈ ;

P₂ is --X(CH₂)_(n) R₈, or --XR₉ Y;

R₃ and R₅ are independently hydrogen, R₁₁, OH, C₁₋₈ alkoxy, S(O)_(q)R₁₁, N(R₆)₂, Br, F, I, Cl, CF₃, NHCOR₆, --R₁₂ CO₂ R₇, --XR₉ --Y or--X(CH₂)_(n) R₈ ;

R₄ is hydrogen, R₁₁, OH, C₁₋₅ alkoxy, S(O)_(q) R₁₁, N(R₆)₂, --X(R₁₁),Br, F, I, Cl or NHCOR₆ wherein the C₁₋₅ alkoxy may be unsubstituted orsubstituted by OH, methoxy or halogen;

R₆ is independently hydrogen or C₁₋₄ alkyl;

R₇ is independently hydrogen, C₁₋₆ alkyl or (CH₂)_(n) Ar;

R₈ is hydrogen, R₁₁, CO₂ R₇, PO₃ H₂, P(O)(OH)R₇, CN, --C(O)N(R₆)₂,tetrazole or OR₆ ;

R₉ is C₁₋₁₀ alkylene, C₂₋₁₀ alkenylene or phenylene all of which may beunsubstituted or substituted by one or more OH, N(R₆)₂, COOH, halogen orXC₁₋₅ alkyl;

R₁₀ is R₃ or R₄ ;

R₁₁ is C₁₋₈ alkyl, C₂₋₈ alkenyl, C₂₋₈ alkynyl all of which may beunsubstituted or substituted by one or more OH, CH₂ OH, N(R₆)₂ orhalogen;

R₁₂ is C₁₋₈ alkylene, C₂₋₈ alkenylene or C₂₋₈ alkenylene;

X is (CH₂)_(n), O, NR₆ or S(O)_(q) ;

Y is CH₃ or --CH₂ X(CH₂)_(n) Ar;

Ar is: ##STR3##

naphythyl, indolyl, pyridyl, thienyl, oxazolidinyl, oxazolyl, thiazolyl,isothiazolyl, pyrazolyl, triazolyl, tetrazolyl, imidazolyl,imidazolidinyl, thiazolidinyl, isoxazolyl, oxadiazolyl, thiadiazolyl,morpholinyl, piperidinyl, piperazinyl, pyrrolyl, or pyrimidyl; all ofwhich may be unsubstituted or substituted by one or more R₃ or R₄groups;

A is C═O, or (C(R₆)₂)_(m) ;

B is --CH₂ -- or --O--;

q is zero, one or two;

n is an integer from () to six;

m is 1, 2 or 3;

and the dotted line indicates the optional presence of a double bond; ora pharmaceutically acceptable salt thereof; provided that when theoptional double bond is present there is no P₁ or R₁₀.

Also included in the invention are pharmaceutically acceptable saltcomplexes.

All alkyl, alkenyl, alkynyl, alkylene, alkenylene, alkenylene, andalkoxy groups may be straight or branched. The term "halogen" is used tomean iodo, fluoro, chloro or bromo. Alkyl groups may be substituted byone or more halogens up to perhalogenation.

The compounds of the present invention may contain one or moreasymmetric carbon atoms and may exist in racemic and optically activeform. All of these compounds and diasteroisomers are contemplated to bewithin the scope of the present invention.

Preferred compounds are those wherein R₁ is X(CH₂)_(n) Ar, (Ar is (a) or(b)), dihydrobenzofuranyl, benzodioxanyl, cyclohexyl, or C₁₋₄ alkyl; R₂is (a), (b), indolyl or hydrogen; R₃ and R₅ are independently hydrogen,OH, C₁₋₅ alkoxy, halogen, R₁₁ CO₂ R₇, C₁₋₄ alkyl, N(R₆)₂, NH(CO)CH₃,--X(CH₂)_(n) R₈, or S(O)_(p) C₁₋₅ alkyl; R₄ is hydrogen, OH, C₁₋₅alkoxy, halogen, C₁₋₄ alkyl, N(R₆)₂, NH(CO)CH₃ or S(O)_(p) C₁₋₅ alkyl;P₁ and P₂ are independently hydrogen, CO₂ H or tetrazole, Ar is (a),(b), or pyridyl; X is (CH₂)_(n) or oxygen and the optional double bondis present.

More preferred are compounds wherein R₁ and R₂ are independently 3,4methylenedioxyphenyl (substituted or unsubstituted by a C₁₋₃ alkoxy orchloro group), phenyl substituted by one or two C₁∝3 alkoxy, O(CH₂)_(n)Ar or O--(CH₂)_(n) C(O) N(H)--SO₂ --Ar groups wherein Ar is phenyl orpyridyl each of which may be substituted by CO₂ H; P₁ is hydrogen, P₂ isCO₂ H; the pyrimidine and pyrizine rings are unsubstituted and thedouble bond is present.

The present invention provides compounds of Formula (Ia-Id) above,##STR4## which can be prepared by a process which comprises: a) forpyrrolo[3,2-b]pyridines (Id) and pyrrolo[2,3-c]pyridines (Ib) in whichthe optional double bond is present and there is no R₁₀ or P₁, reacting(as in this example for pyrrolo[3,2-b]pyridines) a compound of Formula(2), ##STR5## with the appropriate dialkyl oxalate in the presence of abase such as potassium ethoxide in a solvent such as tetrahydrofuran toprovide a nitropyridine of formula (3). ##STR6## Reductive cyclizationof compound (3) in the prescence of a catalyst, such as palladium oncarbon, in a solvent such as ethyl alcohol under an atmosphere ofhydrogen provides a pyrrolopyridine of formula (4) ##STR7## wherein X isC₁₋₅ alkyl. Reacting compound (4) with bromine in a suitable solventsuch as dimethylformamide provides a bromopyrrolopyridine of Formula(5). ##STR8## Coupling of Compound (5) with a boronic acid of formula(6): ##STR9## in the presence of a palladium (0) catalyst, such astetrakis(triphenylphosphine)palladium (0), in a solvent such astoluene/methanol in the presence of a base such as aqueous sodiumcarbonate, at approximately 100° C., provides a pyrrolopyridine ofFormula (7). ##STR10## Aryl boronic acids of Formula (6) may be preparedby transmetallation of aryl halides of Formula (8):

    Ar--Hal                                                    (8)

wherein Hal is Cl, Br or I, with an alkyllithium, such as n-butyllithiumin a solvent such as dry tetrahydrofuran at low temperature (-40° to-78° C.) followed by quenching with a trialkylborate, such astri-isopropylborate, then treatment with an acid such as aqueoushydrochloric.

For compounds in which n is not 0, alkylation of a pyrrolopyridine ofFormula (7) with an halide of Formula (9):

    R.sub.2 --(CH.sub.2).sub.n --Hal                           (9)

in a suitable solvent such as dimethylformamide orhexamethylphosphoramide in the presence of a suitable base such assodium hydride affords compounds of Formula (10), wherein n is not zero.##STR11##

Saponification of esters of Formula (10) with aqueous sodium hydroxidein a solvent such as ethanol or isopropanol at reflux affords compoundsof Formula (11), wherein n is not zero ##STR12## Alternatively,compounds of Formula (7) may be obtained by coupling of compound (5)with an aryl stannane-derivative of Formula (12):

    Ar--SnX.sub.3                                              (12)

in the presence of a palladium (0) catalyst such astetrakis(triphenylphosphine)palladium (0) in a solvent such as dioxan ordimethylformamide at approximately 100° C. in the presence of anhydrouslithium chloride. Aryl stannanes of Formula (10) may be prepared bytransmetallation of aryl halides of Formula (8) with an alkyllithium,such as n-butyllithium, in a solvent such as tetrahydrofuran at lowtemperature (-40° to -78° C.) followed by quenching with atrialkylchlorostannane of Formula (13)

    Cl--SnX.sub.3                                              (13)

b) As an alternative compounds of Formula (5) may be alkylated with anhalide of Formula (9), n#0in a suitable solvent such asdimethylformamide or hexamethylphosphoramide in the presence of asuitable base such as sodium hydride to afford compounds of Formula(14), n is not O. ##STR13## Coupling of Compound (14) with a boronicacid of formula (6) in the presence of a palladium (0) catalyst, such astetrakis(triphenylphosphine)palladium (0), in a solvent such astoluene/methanol in the presence of a base such as aqueous sodiumcarbonate, at approximately 100° C., provides compounds of Formula (10)n is not zero.

As an alternative, compounds of Formula (10), n is not zero, may beobtained by coupling of compound (14) with an aryl stannane derivativeof Formula (12) in the presence of a palladium (0) catalyst such astetrakis(triphenylphosphine)palladium (0) in a solvent such as dioxan ordimethylformamide at approximately 100° C. in the presence of anhydrouslithium chloride.

c) As a further alternative, pyrrolopyridines may be prepared (as inthis example for pyrrolo[2,3-b]pyridines) by a process which comprises:

alkylation of an ester of acetoacetic acid (15) ##STR14## with a halideof Formula (16)

    R.sub.1 --CH.sub.2 Hal                                     (16)

in a suitable solvent such as acetonitrile and a base such as 1,8diazabicyclo[5.4.0]undec-7-ene to afford compounds of Formula (17).Alternatively tetrahydrofuran may be used as the solvent and sodiumhydride as the base for the alkylation. ##STR15## Treatment of acompound of type (17) with an aryl diazonium chloride of Formula (18)##STR16## in a suitable solvent such as ethyl acetate in the presence ofa base such as aqueous sodium hydroxide solution affords, byJapp-Klingemann rearrangement, hydrazones of Formula (19). ##STR17##Treatment of hydrazones of type (19) with a suitable acid such asgaseous hydrogen chloride in a solvent such as ethanol followed byreflux for a period from 0.5 to 12 hours or thermal cyclisation in theabsence of acidic catalysts affords pyrrolo[2,3-b]pyridines of Formula(20) ##STR18## which can be alkylated similarly to compound (7) toprovide compounds of formula (21). ##STR19## d) Compounds of type(Ia-Id) where n=0-6 may be prepared as in this example forpyrrolo[2,3-b]pyridines by a process which comprises:

treatment of a compound of Formula (22) ##STR20## with aqueousformaldehyde solution at reflux affords a product of Formula (23).##STR21## Treatment of compounds of type (23) with aqueous potassiumcyanide at approximately 40°-50° C., affords nitriles of Formula (24).##STR22## Hydrolysis of a nitrile of type (24) with aqueous sodiumhydroxide at reflux followed by acidification with an acid such ashydrochloric affords diacids of Formula (25). ##STR23## Diesterificationof compounds of type (25) is achieved by treatment with a suitable basesuch as 1,8 diazabicyclo[5.4.0]undec-7-ene in a solvent such asacetonitrile or dimethylformamide followed by addition of iodomethane toafford compounds of Formula (26). ##STR24## Diechmann cyclization ofdiesters of type (26) using a base such as sodium methoxide and methanolas solvent at reflux affords products of Formula (27). ##STR25##Treatment of compounds of type (27) with trifluoromethanesulfonicanhydride in pyridine as solvent affords triflates of Formula (28)##STR26## Compounds of Formula (<), X═Me, may be obtained by coupling ofcompound (28) with an aryl stannane derivative of Formula (12) in thepresence of a palladium (0) catalyst such astetrakis(triphenylphosphine)palladium (0) in a solvent such as dioxan ordimethylformamide at approximately 100° C. in the presence of anhydrouslithium chloride. As an alternative compounds of Formula (21), X═Me, canbe prepared by coupling of compound (28) with a boronic acid of formula(6) in the presence of a palladium (0) catalyst, such astetrakis(triphenylphosphine)palladium(0), in a solvent such astoluene/methanol in the presence of a base such as aqueous sodiumcarbonate, at approximately 100° C.

Saponification of compounds of Formula (21), X═Me, to providespyrrolo[2,3-b]pyridines-2-carboxylic acids of Formula (29) can beachieved by treatment with aqueous sodium hydroxide in a solvent such asethanol or isopropanol at reflux. ##STR27##

With appropriate manipulation and protection of any chemicalfunctionalities, synthesis of the remaining compounds of the Formula(Ia-Id) is accomplished by methods analogous to those above and to thosedescribed in the Experimental section.

In order to use a compound of the Formula (Ia-Id) or a pharmaceuticallyacceptable salt thereof for the treatment of humans and other mammals itis normally formulated in accordance with standard pharmaceuticalpractice as a pharmaceutical composition.

Compounds of Formula (Ia-Id) and their pharmaceutically acceptable saltsmay be administered in a standard manner for the treatment of theindicated diseases, for example orally, parenterally, sub-lingually,transdermally, rectally, via inhalation or via buccal administration.

Compounds of Formula (Ia-Id) and their pharmaceutically acceptable saltswhich are active when given orally can be formulated as syrups, tablets,capsules and lozenges. A syrup formulation will generally consist of asuspension or solution of the compound or salt in a liquid carrier forexample, ethanol, peanut oil, olive oil, glycerine or water with aflavouring or colouring agent. Where the composition is in the form of atablet, any pharmaceutical carrier routinely used for preparing solidformulations may be used. Examples of such carriers include magnesiumstearate, terra alba, talc, gelatin, agar, pectin, acacia, stearic acid,starch, lactose and sucrose. Where the composition is in the form of acapsule, any routine encapsulation is suitable, for example using theaforementioned carriers in a hard gelatin capsule shell. Where thecomposition is in the form of a soft gelatin shell capsule anypharmaceutical carrier routinely used for preparing dispersions orsuspensions may be considered, for example aqueous gums, celluloses,silicates or oils and are incorporated in a soft gelatin capsule shell.

Typical parenteral compositions consist of a solution or suspension ofthe compound or salt in a sterile aqueous or non-aqueous carrieroptionally containing a parenterally acceptable oil, for examplepolyethylene glycol, polyvinylpyrrolidone, lecithin, arachis oil, orsesame oil.

Typical compositions for inhalation are in the form of a solution,suspension or emulsion that may be administered as a dry powder or inthe form of an aerosol using a conventional propellent such asdichlorodifluoromethane or trichlorofluoromethane.

A typical suppository formulation comprises a compound of Formula(Ia-Id) or a pharmaceutically acceptable salt thereof which is activewhen administered in this way, with a binding and/or lubricating agent,for example polymeric glycols, gelatins, cocoa-butter or other lowmelting vegetable waxes or fats or their synthetic analogues.

Typical transdermal formulations comprises a conventional aqueous ornon-aqueous vehicle, for example a cream, ointment, lotion or paste orare in the form of a medicated plaster, patch or membrane.

Preferably the composition is in unit dosage form, for example a table,capsule or metered aerosol dose, so that the patient may administer tothemselves a single dose.

Each dosage unit for oral administration contains suitably from 0.1 mgto 500 mg/Kg, and preferably from 1 mg to 100 mg/Kg, and each dosageunit for parenteral administration contains suitably from 0.1 mg to 100mg, of a compound of Formula (Ia-Id) or a pharmaceutically acceptablesalt thereof calculated as the free acid. Each dosage unit forintranasal administration contains suitably 1-400 mg and preferably 10to 200 mg per person. A topical formulation contains suitably 0.01 to1.0% of a compound of Formula (Ia-Id).

The daily dosage regimen for oral administration is suitably about 0.01mg/Kg to 40 mg/Kg, of a compound of Formula (Ia-Id) or apharmaceutically acceptable salt thereof calculated as the free acid.The daily dosage regimen for parenteral administration is suitably about0.001 mg/Kg to 40 mg/Kg, of a compound of the Formula (Ia-Id) or apharmaceutically acceptable salt thereof calculated as the free acid.The daily dosage regimen for intranasal administration and oralinhalation is suitably about 10 to about 500 mg/person. The activeingredient may be administered from 1 to 6 times a day, sufficient toexhibit the desired activity.

No unacceptable toxicological effects are expected when compounds of theinvention are administered to accordance with the present invention.

The biological activity of the compounds of Formula (Ia-Id) aredemonstrated by the following tests:

I. Binding Assay

A) Membrane Preparation (Rat cerebellum or kidney cortex)

Rat cerebellum or kidney cortex were rapidly dissected and frozenimmediately in liquid nitrogen or used fresh. The tissues, 1-2 g forcerebellum or 3-5 g for kidney cortex, were homogenized in 15 mls ofbuffer containing 20 mM Tris HCl and 5 mM EDTA, pH 7.5 at 4° C. using amotor-driven homogenizer. The homogenates were filtered throughcheesecloth and centrifuged at 20,000×g for 10 minutes at 4° C. Thesupernatant was removed and centrifuged at 40,000×g for 30 minutes at 4°C. The resulting pellet was resuspended in a small volume of buffercontaining 50 mM Tris, 10 mM MgCl₂, pH 7.5; aliquotted with small vialsand frozen in liquid nitrogen. The membranes were diluted to give 1 and5 micrograms of protein for each tube for cerebellum and kidney cortexin the binding assay.

Freshly isolated rat mesenteric artery and collateral vascular bed werewashed in ice cold saline (on ice) and lymph nodes were removed fromalong the major vessel. Then, the tissue was homogenized using apolytron in buffer containing 20 nM Tris and 5 mM EDTA, pH 7.5 at 4° C.in 15 ml volume for ˜6 gm of mesenteric artery bed. The homogenate wasstrained through cheesecloth and centrifuged at 2,000 xg for 10 min. at4° C. The supernatant was removed and centrifuged at 40,000 xg for 30min. at 4° C. The resulting pellet was resuspended as explained abovefor cerebellum and kidney cortex. Approximately 10 micrograms ofmembrane protein was used for each tube in binding experiments.

B) CHO Cell Membrane Preparation

CHO cells stably transfected with human ET_(A) and ET_(B) receptors weregrown in 245 mm×245 mm tissue culture plates in Dulbecco's modifiedEagle's medium (DMEM) supplemented with 10% fetal bovine serum (FBS).The confluent cells were washed with DPBS (Dulbecco's phosphate bufferedsaline) containing protease inhibitor cocktail (5 mM EDTA, 0.5 mM PMSF,5 ug/ml leupeptin, and 0.1 U/ml aprotinin) and scraped in the samebuffer. After centrifugation at 800 xg, the cells were lysed by freezingin liquid nitrogen and thawing on ice followed by homogenization (30times using glass dounce homogenizer) in lysis buffer containing 20 mMTris HCl, pH 7.5 and the protease inhibitor cocktail. After an initialcentrifugation at 800 xg for 10 min to remove unbroken cells and nuclei,the supernatants were centrifuged at 40,000×g for 15 min and the pelletwas resuspended in 50 mM Tris HCl, pH 7.5 and 10 mM MgCl₂ and stored insmall aliquots at -70° C. after freezing in liquid N₂. Protein wasdetermined using BCA method and bovine serum albumin as the standard.

C) [¹²⁵ I]ET-1 Binding Protocol

[¹²⁵ I]ET-1 binding to membranes from rat cerebellum (2-5 mgprotein/assay tube) or kidney cortex (3-8 micrograms protein/assay tube)or CHO cell membranes (containing 4-6 and 1-2 micrograms of membraneprotein for ET_(A) and ET_(B) receptors, respectively) were measuredafter 60 minutes incubation at 30° C. in 50 mM Tris HCl, 10 mM MgCl₂,0.05% BSA, pH 7.5 buffer in a total volume of 100 microliters. Membraneprotein was added to tubes containing either buffer or indicatedconcentration of compounds. [¹²⁵ I]ET-1 (2200 Ci/mmol) was diluted inthe same buffer containing BSA to give a final concentration of 0.2-0.5nM ET-1. Total and nonspecific binding were measured in the absence andpresence of 100 nM unlabelled ET-1. After the incubation, the reactionswere stopped with 3.0 ml cold buffer containing 50 mM Tris and 10 mMMgCl₂, pH 7.5. Membrane bound radioactivity was separated from freeligand by filtering through Whatman GF/C filter paper and washing thefilters 5 times with 3 ml of cold buffer using a Brandel cell harvester.Filter papers were counted in a gamma counter with an efficiency of 75%.IC₅₀ 's for the compounds of this invention range from 0.01 nm to 50 uM.

II. In Vitro Vascular Smooth Muscle Activity

Rat aorta are cleaned of connective tissue and adherent fat, and cutinto ring segments approximately 3 to 4 mm in length. Vascular rings aresuspended in organ bath chambers (10 ml) containing Krebs-biscarbonatesolution of the following composition (millimolar): NaCl, 112,0; KCl,4.7; KH₂ PO₄, 1.2; MgSO₄, 1.2; CaCl₂, 2.5; NaHCO₃, 25.0; and dextrose,11.0. Tissue bath solutions are maintained at 37° C. and aeratedcontinuously with 95% O₂ /5% CO₂. Resting tensions of aorta aremaintained at 1 g and allowed to equilibrate for 2 hrs., during whichtime the bathing solution is changed every 15 to 20 min. Isometrictensions are recorded on Beckman R-611 kynographs with Grass FT03force-displacement transducer. Cumulative concentration-response curvesto ET-1 or other contractile agonists are constructed by the method ofstep-wise addition of the agonist. ET-1 concentrations are increasedonly after the previous concentration produces a steady-statecontractile response. Only one concentration-response curve to ET-1 isgenerated in each tissue. ET receptor antagonists are added to pairedtissues 30 min prior to the initiation of the concentration-response tocontractile agonists.

ET-1 induced vascular contractions are expressed as a percentage of theresponse elicited by 60 mM KCl for each individual tissue which isdetermined at the beginning of each experiment. Data are expressed asthe mean ±S.E.M. Dissociation constants (K_(b)) of competitiveantagonists were determined by the standard method of Arunlakshana andSchild. The potency range for compounds of this invention range from 0.1nM to 50 mm.

The following examples are illustrative and are not limiting of thecompounds of this invention.

EXAMPLE 13-(4-Methoxyphenyl)-1-(3,4-methylenedioxy-phenylmethyl)pyrrolo[2,3-b]pyridine-2-carboxylicacid

a) Ethyl2-(4-methoxybenzyl)-3-oxobutyrate

A solution of ethyl acetoacetate and 4-methoxybenzyl chloride is stirredunder an argon atmosphere with 1,8-diazabicyclo[5.4.0]undec-7-ene atroom temperature in CH₃ CN. The mixture is partitioned between 3 N HCland EtOAc. The organic extract is washed successively with H₂ O, aqueousNaHCO₃, H₂ O and saturated aqueous NaCl and dried (Na₂ SO₄). The solventis removed in vacuo to afford the title compound.

b) Ethyl 3-(4-methoxyphenyl)pyrrolo[2,3-b]pyridine-2-carboxylate

To a solution of ethyl 2-(4-methoxybenzyl)-3-oxobutyrate in EtOAcstirred at ice bath temperature under an argon atmosphere is added anaquious solution of NaOH. This is immediately followed by the additionof an aqueous solution of pyrid-2-yldiazonium chloride [prepared from2-aminopyridine in 6 N HCl and NaNO₂ ]. The mixture is partitionedbetween EtOAc and H₂ O. The aqueous layer is washed with EtOAc. Thecombined organic extracts are washed with saturated aqueous NaClsolution, dried (Na₂ SO₄) and the solvent is removed in vacuo. Theresidue is dissolved in EtOH and the solution is saturated with HCl gas.This is refluxed then cooled to room temperature and partitioned EtOAcand saturated aqueous NaHCO₃ solution. The aqueous layer is washed withEtOAc. The combined organic extract is washed with H₂ O then saturatedaqueous NaCl solution, dried (Na₂ SO₄) and the solvent is removed invacuo. The residue is purified by chromatography to afford the titlecompound.

c) Ethyl3(4-methoxyphenyl)-1-(3,4-methylenedioxybenzyl)pyrrolo[2,3-b]pyridine-2-carboxylate

To a solution of ethyl3-(4-methoxyphenyl)pyrrolo[2,3-b]pyridine-2-carboxylate in HMPA stirredat ice bath temperature under an argon atmosphere is added NaH. Asolution of piperonyl chloride in HMPA is added and the ice bathremoved. The reaction mixture is stirred at room temperature thenpartitioned between 3 N HCl and EtOAc. The organic extract is washedsuccessively with H₂ O, aqueous NaHCO₃, H₂ O and saturated aqueous NaCland dried (Na₂ SO₄). The solvent is removed in vacuo. The residue ispurified by chromatography to afford the title compound.

d)3-(4-Methoxyphenyl)-1-(3,4-methylenedioxybenzyl)-pyrrolo[2,3-b]pyridine-2-carboxylicacid

A solution of ethyl3-(4-methoxyphenyl)-1-(3,4-methylenedioxybenzyl)-pyrrolo[2,3-b]pyridine-2-carboxylatein EtOH with aqueous 1 N NaOH is stirred under an argon atmosphere firstat room temperature then at reflux temperature. The reaction mixture iscooled to room temperature then poured into H₂ O and the solvent volumereduced in vacuo. The aqueous solution is extracted with Et₂ O and theEt₂ O extract discarded. The aqueous layer is acidified with 6 N HCl andthe product extracted into EtOAc. The organic extract is washed with H₂O then saturated aqueous NaCl, dried (Na₂ SO₄) and the solvent removedin vacuo to afford the title compound.

EXAMPLE 23-[2(2-carboxyphenylmethoxy)-4-methoxyphenyl]-1-(3,4-methylenedioxybenzyl)pyrrolo[2,3-b]pyridine-2-carboxylicacid EXAMPLE 3 3-[4-Methoxy-2-(N-phenylsulfony)carboxamidomethoxy)phenyl]-1-(3,4-methylenedioxybenzyl)pyrrolo[2,3-b]pyridine-2-carboxylicacid EXAMPLE 41-[(2-Carboxymethoxy-4-methoxyphenyl)methyl]-3-(3,4-methylenedioxyphenyl)pyrrolo[2,3-b]pyridine-2-carboxylicacid m.p. 244-255 EXAMPLE 5

Formulations for pharmaceutical use incorporating compounds of thepresent invention can be prepared in various forms and with numerousexcipients. Examples of such formulations are given below.

Inhalant Formulation

A compound of formula Ia, Ib, Ic, or Id (1 mg to 100 mg) is aerosolizedfrom a meterred dose inhaler to deliver the desired amount of drug peruse.

    ______________________________________                                        Tablets/Ingredients  Per Tablet                                               ______________________________________                                        1.      Active ingredient                                                                              40 mg                                                        (Cpd of Form. Ia, Ib, Ic or Id)                                       2.      Corn Starch      20 mg                                                3.      Alginic acid     20 mg                                                4.      Sodium alginate  20 mg                                                5.      Mg stearate      1.3 mg                                                                        2.3 mg                                               ______________________________________                                        Procedure for tablets:                                                        ______________________________________                                        Step 1                                                                              Blend ingredients No. 1, No. 2, No. 3 and No. 4 in a suitable                  mixer/blender.                                                         Step 2                                                                              Add sufficient water portion-wise to the blend from Step 1 with               careful mixing after each addition. Such additions of water and               mixing until the mass is of a consistency to permit its conversion            to wet granules.                                                        Step 3                                                                              The wet mass is converted to granules by passing it through an                oscillating granulator using a No. 8 mesh (2.38 mm) screen.             Step 4                                                                              The wet granules are then dried in an oven at 140° F.                  (60° C.) until dry.                                              Step 5                                                                              The dry granules are lubricated with ingredient No. 5.                  Step 6                                                                              The lubricated granules are compressed on a suitable tablet                   press.                                                                  ______________________________________                                    

Parenteral Formulation

A pharmaceutical composition for parenteral administration is preparedby dissolving an appropriate amount of a compound of formula Ia, Ib, Icor Id in polyethylene glycol with heating. This solution is then dilutedwith water for injections Ph Eur. (to 100 ml). The solution is thensteriled by filtration through a 0.22 micron membrane filter and sealedin sterile containers.

We claim:
 1. A compound of the formula Ia ##STR28## wherein: R₁ is--X(CH₂)_(n) Ar or cyclohexyl;R₂ is a moiety of formula (a) or (b) orindolyl; P₁ is CO₂ H or tetrazole; P₂ is hydrogen; R₃ and R₅ areindependently hydrogen, R₁₁, OH, C₁₋₈ alkoxy, S(O)_(q) R₁₁, N(R₆)₂, Br,F, I, Cl, CF₃, NHCOR₆, --R₁₂ CO₂ R₇, --XR₉ --Y or --X(CH₂)_(n) R₈ ; R₄is hydrogen, R₁₁, OH, C₁₋₅ alkoxy, S(O)_(q) R₁₁,N(R₆)₂, --X(R₁₁), Br,F,I, Cl or NHCOR₆ wherein the C₁₋₅ alkoxy may be unsubstituted orsubstituted by OH, methoxy or halogen; R₆ is independently hydrogen orC₁₋₄ alkyl; R₇ is independently hydrogen, C₁₋₆ alkyl or (CH₂)_(n) Ar; R₈is hydrogen, R₁₁, CO₂ R₇, PO₃ H₂, P(O)(OH)R₇, CN, --C(O)N(R₆)₂,tetrazole or OR₆ ; R₉ is C₁₋₁₀ alkylene, C₂₋₁₀ alkenylene or phenyleneall of which may be unsubstituted or substituted by one or more OH,N(R₆)₂, COOH, halogen or XC₁₋₅ alkyl; R₁₀ is R₃ or R₄ ; R₁₁ is C₁₋₈alkyl, C₂₋₈ alkenyl, C₂₋₈ alkynyl all of which may be unsubstituted orsubstituted by one or more OH, CH₂ OH, N(R₆)₂ or halogen; R₁₂ is C₁₋₈alkylene, C₂₋₈ alkenylene or C₂₋₈ alkylnylene; X is (CH₂)_(n), or O,; Yis CH₃ or --CH₂ X(CH₂)_(n) Ar; Ar is: ##STR29## or naphthyl, indolyl,pyridyl, thienyl, oxazolidinyl, oxazolyl, thiazolyl, isothiazolyl,pyrazolyl, triazolyl, tetrazolyl, imidazolyl, imidazolidinyl,thiazolidinyl, isoxazolyl, oxadiazolyl, thiadiazolyl, morpholinyl,piperidinyl, piperazinyl, pyrrolyl, or pyrimidyl; all of which may beunsubstituted or substituted by one or more R₃ or R₄ groups: A is C═O,or (C(R₆)₂)_(m) ; B is --CH₂ -- or --O--; q is zero, one or two; n is aninteger from 0 to six; m is 1, 2 or 3; and the dotted line indicates theoptional presence of a double bond; or a pharmaceutically acceptablesalt thereof; provided that when the optional double bond is presentthere is no P₁ or R₁₀.
 2. A compound of claim 1 wherein R₃ R₅ areindependently hydrogen, OH, C₁₋₅ alkoxy, halogen, R₁₁ CO₂ R₇, C₁₋₄alkyl, N(R₆)₂, NH(CO)CH₃, --X(CH₂)_(n) R₈, phenyl or S(O)_(p) C₁₋₅alkyl; R₄ is hydrogen, OH, C₁₋₅ alkoxy, halogen, C₁₋₄ alkyl, N(R₆)₂,NH(CO)CH₃ or S(O)_(p) C₁₋₅ alkyl; Ar is (a), (b) or pyridyl; X is(CH₂)_(n) or oxygen and the optional double bond is present.
 3. Acompound of claim 1 wherein R is dihydrobenzofuranyl, benzodioxanyl orcyclhexyl.
 4. A pharmaceutical composition comprising a compound ofclaim 1 and a pharmaceutically acceptable carrier.
 5. A method oftreatment of diseases caused by an excess of endothelin which comprisesadministering to a subject in need thereof, an effective amount toantagonize endothelin receptors of a compound of claim
 1. 6. A method oftreating hypertension which comprises administering to a subject in needthereof an effective amount of a compound of claim
 1. 7. A method oftreating renal failure which comprises administering to a subject inneed thereof, an effective amount of a compound of claim
 1. 8. A methodof treating cerebrovascular disease which comprises administering to asubject in need thereof, an effective amount of a compound of claim 1.